Our development of the isolated axoplasm preparation from the squid giant axon has provided a valuable paradigm for dissecting the molecular mechanisms of fast axonal transport. For the first time, it has been possible to demonstrate that fast axonal transport involves a novel mechanism for motility in cells that is not based on either actomyosin or dynein. The use of a nonhydrolyzable ATP analogue, adenylyl imidodiphosphate (AMP-PNP), as a pharmacological probe has permitted identification of a novel brain ATPase activity with the properties predicted for the fast axonal transport motor. The studies proposed in this application are designed to characterize the ATPase associated with the motor for fast transport; to determine the cellular and subcellular distribution of the motor; and to evaluate possible mechanisms for regulation of fast axonal transport. The proposed experiments include further purification of the ATPase, immunochemical analyses of ATPase function and distribution, kinetic studies of ATPase activity, analyses of interactions with vesicles and microtubules, and evaluation of the molecular bases of motility and directionality. These studies will provide the first detailed information about the molecular mechanisms of fast axonal transport and the properties of the motor for vesicle movements in neurons and other cells.